Francisella tularensis is a Gram-negative, facultative intracellular pathogen that causes tularemia, a debilitating and potentially fatal disease that affects humans and a wide range of animals. Infections can be acquired through bites from an arthropod vector, skin lesions, ingestion of contaminated food or water, and by inhalation of as few as 10 bacteria (Dennis et al., JAMA 285:2763-73, 2001). The low dose required to cause tularemia by aerosol route resulted in the development of F. tularensis for use as a biological weapon by several national weapons programs. The U.S. Centers for Disease Control and Prevention (CDC) classified F. tularensis as a Category A bioterrorism agent, members of which are considered most serious in posing a risk to national security. There is currently no approved vaccine available in the U.S. or Europe. Thus, the development of a vaccine against F. tularensis is an international priority.
Although the molecular mechanisms of F. tularensis pathogenesis remain obscure, replication in human and animal macrophages is central to the organism's ability to cause tularemia (Fortier et al., Immunol. Ser. 60:349-61, 1994). Several F. tularensis genes associated with intracellular growth have been identified, including iglB, iglC, mglA, pdpD, and a clpB homolog (Baron and Nano. Mol. Microbiol. 29:47-259, 1998; Golovliov et al., FEMS Microbiol. Lett. 222:273-80, 2003; Gray et al., FEMS Microbiol. Lett. 215:53-6, 2002; Lai et al., Microb. Pathog. 37:225-30, 2004; and Lauriano et al., Proc. Natl. Acad. Sci. USA 101:4246-9, 2004). Although many of the genes in the F. tularensis pathogenicity island (FPI) have been proposed to contribute to its survival and growth in macrophages (Larsson et al., Nat. Genet. 37:153-9, 2005; Nano et al., J. Bacteriol. 186:6430-6, 2004) none have arisen as potential vaccine candidates.
Four main subspecies of F. tularensis are commonly recognized: tularensis (type A), holarctica (type B), novicida, and mediasiatica. All of these biotypes share greater than 95% DNA sequence identity (Broekhuijsen et al., J. Clin. Microbiol. 41:2924-31, 2003). Although type A and type B strains are highly infectious, only type A strains cause significant mortality in humans. The current live vaccine strain (LVS) is an attenuated type B strain that provides varying levels of protection against challenge with type A F. tularensis strains (Chen, et al., Microb. Patholg. 36:311-8, 2004; Chen et al., Vaccine 21:3690-700, 2003; Conlan et al., Vaccine 23:2477-85, 2005; Green et al., Vaccine 23:2680-6, 2005; Shen, et al., Vaccine 22:2116-21, 2004; Wu et al., Infect. Immun. 73:2644-54, 2005). However, several limitations prevent the licensing of this vaccine. For example, the genetic basis of LVS attenuation and protection remains unknown. In addition, culturing LVS under certain conditions can lead to poorly immunogenic colony variants, demonstrating this organism's genetic instability (Cowley et al., Mol. Microbiol. 20:867-74, 1996; Eigelsbach and Downs. J. Immunol. 87:415-25, 1961). Also, this vaccine does not confer protection to all vaccinated subjects (McCrumb, Bacteriol. Rev. 25:262-7, 1961; Saslaw et al., Arch. Intern. Med. 107:702-14, 1961). Furthermore, LVS protection against aerosol challenge is variable and depends on the route of immunization as well as the host (Chen, et al., Microb. Patholg. 36:311-8, 2004; Chen et al., Vaccine 21:3690-700, 2003; Conlan et al., Vaccine 23:2477-85, 2005; and Shen et al., Vaccine 22:2116-21, 2004). This last point is relevant when considering F. tularensis as a biological weapon, as aerosol dispersal is the most likely route of delivery. These limitations demonstrate the need for an approved tularemia vaccine.